The proposed research will study the mechanisms by which hypothalamic substances regulate the secretion of prolactin from the GH3 anterior pituitary cell line. The hypothalamic substances to be studied are thyrotropin releasing hormone (TRH) which increases prolactin secretion and dopamine (DA) and somatostatin which decrease prolactin secretion. A combination of electrophysiological and biochemical techniques will be used to analyze the membrane transducing action of these substances and the role of Ca++ ions in the modulation of secretion by these substances. The passive and voltage dependent properties of the GH3 cell membrane, and the effect of TRH, DA and somatostatin on these properties will be analyzed by current clamp, voltage clamp and single channel-patch clamp analysis. The effects of these compounds on Ca++ ion homeostasis will be analyzed by 45Ca flux studies and measurements of intracellular Ca++ ion levels with a Ca++ sensitive fluorescent dye and a Ca++ sensitive microelectrode. The effects of these substances on the control of exocytosis by intracellular Ca++ will be analyzed by using cells permeabilized by high voltage dielectric breakdown or by digitonin disruption of the plasma membrane. Information gained from these studies are important for our basic understanding of the regulation of hormone secretion from endocrine cells. In addition such information may provide clues for the possible therapeutic control of hormone secretion from endocrine tumors or from disorders of hormone regulation involving failure of a particular endocrine cell type to secrete hormone.